Purification of Plasmodium Sporozoites Enhances Parasite-Specific CD8+ T Cell Responses

Abstract

Malaria infection caused by Plasmodium parasites continues to cause enormous morbidity and mortality in areas where it is endemic, and there is no licensed vaccine capable of inducing sterile protection. Hyperimmunization with attenuated whole sporozoites can induce sterile protective immune responses targeting preerythrocytic antigens. Most animal models of hyperimmunization rely on sporozoites dissected from mosquito salivary glands and injected without further purification. In BALB/c mice, repeated small doses of P. yoelii sporozoites progressively expand the population of sporozoite-specific CD8(+) T cells. In this study, large secondary doses of unpurified sporozoites unexpectedly led to contraction of sporozoite-specific CD8(+) T cell responses in sporozoite-primed mice. While sporozoite-primed CD8(+) T cells alternatively can be expanded by secondary exposure to Listeria monocytogenes expressing recombinant Plasmodium antigens, such expansion was potently inhibited by coinjection of large doses of unpurified sporozoites and by uninfected salivary glands alone. Purification of sporozoites away from mosquito salivary gland debris by density gradient centrifugation eliminated salivary gland-associated inhibition. Thus, the inhibitory effect appears to be due to exposure to uninfected mosquito salivary glands rather than sporozoites. To further assess the effect of salivary gland exposure on later sporozoite vaccinations, mice were immunized with uninfected salivary glands from a single mosquito. Compared to naive mice, salivary gland presensitization reduced subsequent liver burdens by 71%. These data show that a component(s) in mosquito salivary glands reduces liver infection, thereby limiting antigen dose and contributing to lower-magnitude T cell responses. These findings suggest that sporozoite immunogenicity studies be performed using purified sporozoites whenever feasible.